4MJO

Human liver fructose-1,6-bisphosphatase(d-fructose-1,6-bisphosphate, 1-phosphohydrolase) (e.c.3.1.3.11) complexed with the allosteric inhibitor 3

Summary for 4MJO

• Entry DOI: 10.2210/pdb4mjo/pdb
• Related: 2jjk 2vt5
• Descriptor: Fructose-1,6-bisphosphatase 1, N-({4-bromo-6-[(methylcarbamoyl)amino]pyridin-2-yl}carbamoyl)-5-(2-methoxyethyl)-4-methylthiophene-2-sulfonamide (3 entities in total)
• Functional Keywords: allostery, hydrolase (phosphoric monoester), hydrolase-hydrolase inhibitor complex, hydrolase/hydrolase inhibitor
• Biological source: Homo sapiens (human)
• Total number of polymer chains: 8
• Total formula weight: 299150.63

Authors

Ruf, A.,Joseph, C.,Tetaz, T.,Benz, J. (deposition date: 2013-09-04, release date: 2013-11-06, Last modification date: 2023-09-20)

Primary citation

Cubrilovic, D.,Haap, W.,Barylyuk, K.,Ruf, A.,Badertscher, M.,Gubler, M.,Tetaz, T.,Joseph, C.,Benz, J.,Zenobi, R.
Determination of protein-ligand binding constants of a cooperatively regulated tetrameric enzyme using electrospray mass spectrometry.
Acs Chem.Biol., 9:218-226, 2014

PubMed Abstract: This study highlights the benefits of nano electrospray ionization mass spectrometry (nanoESI-MS) as a fast and label-free method not only for determination of dissociation constants (KD) of a cooperatively regulated enzyme but also to better understand the mechanism of enzymatic cooperativity of multimeric proteins. We present an approach to investigate the allosteric mechanism in the binding of inhibitors to the homotetrameric enzyme fructose 1,6-bisphosphatase (FBPase), a potential therapeutic target for glucose control in type 2 diabetes. A series of inhibitors binding at an allosteric site of FBPase were investigated to determine their KDs by nanoESI-MS. The KDs determined by ESI-MS correlate very well with IC50 values in solution. The Hill coefficients derived from nanoESI-MS suggest positive cooperativity. From single-point measurements we could obtain information on relative potency, stoichiometry, conformational changes, and mechanism of cooperativity. A new X-ray crystal structure of FBPase tetramer binding ligand 3 in a 4:4 stoichiometry is also reported. NanoESI-MS-based results match the current understanding of the investigated system and are in agreement with the X-ray structural data, but provide additional mechanistic insight on the ligand binding, due to the better dynamic resolution. This method offers a powerful approach for studying other proteins with allosteric binding sites, as well.

PubMed: 24128068
DOI: 10.1021/cb4007002

Experimental method

X-RAY DIFFRACTION (2.4 Å)